The present invention relates to a novel N-terminated peptide antibody of the profragment of human prorenin, referred to as "pf" hereinafter, capable of forming an immune complex by combining with human prorenin, a renin-active substance formed when the antibody is combined with human prorenin and an assay reagent for prorenin by using the same.
Prorenin is a substance having no enzyme activity, which is produced mainly in the kidney as a precursor of the renin of the completely matured type or, namely, as the renin of the completely matured type combined with a profragment consisting of 43 amino acid residues. It is known that, when diabetes causes vascular disorders, the concentration of human prorenin in blood is increased depending on the seriousness of the disease while the concentration is decreased along with alleviation of the disease. It is proposed accordingly to take the concentration of human prorenin in blood as a marker of the diabetic vascular disorder (see The New England Journal of Medicine, volume 312, 1985, pages 1412-1417, Memoirs of Tokyo Women's Medical College, volume 60, 1990, pages 342-350, and Clinical Investigator, volume 71, 1993, pages 3-6).
In connection therewith, several proposals have been made heretofore for the assay of the prorenin concentration in human plasma. A problem in these prior art methods is that, because human plasma contains both of the renin of the completely matured type as an enzymatic protein and the enzymatically inactive human prorenin as a precursor of the renin of the completely matured type, an indirect method must be undertaken in which the human prorenin in plasma is first partially activated with an acid at a low temperature and then converted into the renin of the completely matured type by using trypsin followed by the determination of the overall amount of renin by an enzymological method as the overall amount of renin activity or by an immunological method as the overall amount of active renin and the amount of the human prorenin is calculated as a difference obtained by subtracting the separately obtained value of the renin activity by an enzymological method or the amount of active renin by an immunological method from the above obtained overall renin activity or overall amount of active renin.
The "renin of the completely matured type" here implied is a structural body derived from prorenin after separation of the profragment part therefrom by a processing enzyme and can exhibit the renin activity since the enzymatically active part is open. The renin activity above mentioned is the enzymatic activity as an inherent function of the renin of the completely matured type which is an enzyme protein or, namely, the activity to produce angiotensin (Ang I) by selectively acting on the renin substrate (angiotensinogen).
A discovery was recently made to convert inactive human prorenin into an open-type structure by utilizing the effect of bonding thereof with a low-molecular renin inhibitor and, as a result of this discovery, a possibility has been established of an immunological method for the assay of the overall renin activity by utilizing a monoclonal antibody capable of specifically recognizing the proximity of the renin active part and a monoclonal antibody capable of recognizing both of the human prorenin and the renin of the completely matured type along with development of a method in which the amount of active renin is determined without addition of a renin inhibitor to calculate the amount of human renin from the difference between the overall active renin and the active renin determined as above (see Clinical Chemistry, volume 42, 1996, pages 1051-1063). As compared with the prior art method for the trypsin-activated enzymological method or the trypsin activation method for the assay of renin activity, this method, though advantageous in respect of the possibility of suppressing transient decomposition of the human prorenin and renin of the completely matured type by trypsin as well as in the good correlation with the trypsin activation method, has a defect that the renin activity obtained thereby is positively biased as compared with the true renin activity for diabetic cases where an increase is noted in the prorenin titer in blood. Moreover, this method is troublesome because the assay must be conducted each time for the renin activity and for the overall renin activity.
Besides, an immunological method of assay is known which is a direct method by utilizing a monoclonal antibody capable of recognizing the C-terminal of the pf in the human prorenin or, namely, the 29th to 43rd amino acid residues as an antigen and a renin monoclonal antibody capable of recognizing both of the human prorenin and renin of the completely matured type (see Journal of Clinical Endocrinology and Metabolism, volume 75, 1992, pages 617-623).
This method is advantageous because the method has, in addition to the very good correlation with the trypsin activation method, a sensitivity so high that the method can be used even for the assay of the prorenin titer in blood taken from a healthy person. On the other hand, the method has defects in that, as a trend, the value obtained by the assay is negatively biased to be as low as about 80% as compared with the value obtained by the trypsin activation method, this trend being particularly remarkable for a patient of a disease giving an increased human prorenin titer, along with the indefiniteness of the part where the renin monoclonal antibody is recognized.